The present invention relates to a process for preparing a heteropolysaccharide by cultivating a bacterium, to bacteria having the characteristic of increased heteropolysaccharide production, also to a method of producing such bacteria.
It is known that a heteropolysaccharide (in particular, a succinoglucan) can be prepared by subjecting a carbohydrate source to fermentation by certain micro-organisms such as Pseudomonas sp. NCIB 11592 as described in EP-A-0040445 (K 1480 EPC). That specification gives a full description of the use of heteropolysaccharides as a viscosifier for aqueous solutions used in oil recovery. More recently, micro-organism NCIB 11883, as described by Linton et al (1987)(this and other references by author and ear are hereinafter defined), and in EP-A-0138255 (K 1924 EPC), has been isolated; it appears to produce polysaccharide at a considerably faster rate than NCIB 11592. Further, the productivity in terms of viscosifying power as expressed by the dilution factor of the culture broth, is considerably higher in strain NCIB 11883 than in Pseudomonas sp. NCIB 11592.
It appears that most bacteria capable of growing anaerobically possess phosphotransferase systems (PTS) that catalyze the uptake and concomitant phosphorylation of glucose at the expense of phosphoenolpyruvate. By contrast, this type of glucose uptake system appears to be absent from the very limited number of aerobic bacteria in which glucose transport has been studied, although a fructose-specific PTS has been identified in Pseudomonas aeruginosa.
Two alternative routes for glucose uptake, both involving periplasmic proteins, have been identified in P. aeruginosa. During growth in continuous culture under glucose excess, oxygen-sufficient conditions this organism does not take up glucose directly but produces two periplasmic enzymes, glucose dehydrogenase and gluconate dehydrogenase, that sequentially oxidise glucose to gluconate and 2-ketogluconate which are then taken up via specific transport systems (Midgley and Dawes, 1973; Roberts et al, 1973; Whiting et al, 1973). This extracellular oxidation system is repressed during glucose-limited growth (Whiting et al, 1976), and glucose is taken up directly via a high-affinity transport system (Midgley and Dawes, 1973). The latter involves a periplasmic, glucose-binding protein (Stinson et al, 1977), and thus appears to be analogous to the binding protein-dependent transport systems for certain sugars, amino-acids and inorganic ions that have been investigated in detail in enteric bacteria (Ames, 1986).
Agrobacterium radiobacter NCIB 11883, the recently-isolated aerobic bacterium of industrial importance, is unable to metabolize glucose using the indirect, oxidative route since it produces a non-functional, apoenzyme form of glucose dehydrogenase that lacks the PQQ (pyrroloquinoline quinone) co-factor (Linton et al, 1986; Cornish et al, 1987).